The goal of this Phase I project is to evaluate 2-adamantanamine compounds as a potential therapeutic to treat recurrent oropharyngeal candidiasis (OPC) in HIV patients. 2-adamantanamine is a close structural analog of the antivirals amantadine and rimantadine, making it an attractive drug candidate. Refractory OPC is currently reported in 4-5% of HIV infected individuals. Upon adherence to a surface, C. albicans forms invasive hyphae. We found that 2-adamantanamine prevented invasion and hyphal elongation, two key events in Candida pathogenesis. Recalcitrance to antimicrobial treatment is another important feature of OPC. Apart from forming hyphae, attachment leads to production of drug-tolerant persister cells (LaFleur et al., 2006). We find that patients with unresolved OPC carry high-persistent (hip) mutants of C. albicans, apparently selected by antimicrobial therapy. A combination of AC17 and miconazole was able to eradicate C. albicans cells, including persisters in hip strains. We will evaluate the ability of 2-adamantanamine compounds alone and in combination with miconazole to suppress C. albicans in a reconstituted human oral epithelium model of candidiasis. This will serve as a basis for a Phase II drug development project. The Specific Aims of the project are: Aim 1. In vitro validation of 2-adamantanamine compounds. Many 2-adamantanamine analogs are available commercially, facilitating the evaluation of this class of compounds. The efficacy of 2- adamantanamine and its analogs in inhibiting invasion and hyphal elongation of Candida albicans clinical isolates in vitro will be measured. 26 available analogs will be examined, and 3 of the most potent compounds will be tested against a panel of 20 clinical isolates in order to determine the minimal concentration inhibiting hyphal elongation and invasion in 90% of the strains (MHE90). Milestone: Identify three 2-adamantanamine analogs with a satisfactory MHE90. Aim 2. Efficacy of 2-adamantanamine analogs in a reconstituted human oral epithelium model of candidiasis. The efficacy of the selected 2-adamantanamine analogs against Candida will be evaluated in the reconstituted human oral epithelium model. Compounds will be tested prophylactically and as a treatment for established infection alone and in combination with miconazole and clotrimazole. Compound efficacy will be measured by colony count, LDH release by the epithelium, and microscopically. Milestone: For adamantanamine stand-alone studies: significant reduction in infection. In combination studies, significant reduction in pathogen burden compared to miconazole and clotrimazole alone. Aim 3. Target and mechanism of action of 2-adamantanamine. Identification of resistance mutations by whole genome sequencing, and drug profiling with DNA arrays will be used to determine the target of 2- adamantanamine. Once candidate targets are identified, they will be validated by constructing strains with decreased vs. increased expression of the protein in Candida. We expect increased susceptibility of a strain diminished in target expression as compared to a strain overexpressing the target. Milestone: Identify cellular target(s) responsible for preventing invasion and potentiating miconazole in Candida. )